Target Proteins for Linkages in Membranes of Hair Cells

毛细胞膜中连接的靶蛋白

基本信息

批准号：
6626306
负责人：
GLEN M WATSON
金额：
$ 6.23万
依托单位：
UNIVERSITY OF LOUISIANA AT LAFAYETTE
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-04-22 至 2005-03-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Hair bundle mechanoreceptors detect sound, motion, acceleration and gravity. They are missing, damaged or nonfunctioning in millions of Americans suffering from hearing or balance disorders. Hair bundles are composed of numerous stereocilia interconnected by extracellular linkages. Whereas most of the linkages may tie the hair bundle together so that it behaves as a single, mechanical unit, certain linkages may function as "gating springs" that open transduction channels. Hair bundle mechanoreceptors on tentacles of sea anemones are similar to those of the acousticolateralis system except that anemone hair bundles self-repair following extensive trauma. Repair is accomplished by "repair proteins" that apparently include replacement linkages for those lost during trauma. Exogenously supplied repair proteins rapidly restore structural integrity and function to traumatized hair bundles. According to the primary hypothesis of this proposal, extracellular linkages are not themselves integral proteins, but instead are attached to integral proteins called, "membrane target proteins." In this proposal, a specific chromatographic fraction of the repair protein mixture will be used as an affinity trap to isolate candidate membrane target proteins. These proteins will be blotted to PVDF membrane and partially sequenced. The resulting information will be used to synthesize peptides that will be used as immunogens. Peptide-specific Antibodies are expected to label specific domains of hair bundles. A cDNA library to tentacle tissue will be constructed and then used to transfect bacteria. Recombinant clones will be screened using the peptide-specific antibodies. Single stranded DNA will be isolated from immunopositive clones and then sequenced. This research may lead to the isolation of the transduction channel among other important membrane proteins in hair cells. This information will likely lead to a better understanding of the molecular basis of certain forms of deafness and balance disorders.

描述（由申请人提供）：发束机械感受器检测声音，运动、加速度和重力。它们丢失、损坏或无法工作数百万美国人患有听力或平衡障碍。发束由许多通过细胞外连接相互连接的静纤毛组成。然而大多数连接可能会将发束绑在一起，使其表现出作为一个单一的机械单元，某些连杆可以起到“门控弹簧”的作用打开转导通道。触手上的毛束机械感受器海葵与声外侧系统相似，除了海葵毛束在大面积创伤后能够自我修复。修复是通过显然包含替换连接的“修复蛋白”来完成对于那些在创伤中失去的人。快速提供外源修复蛋白恢复受损发束的结构完整性和功能。根据根据该提议的主要假设，细胞外联系不是本身是完整蛋白质，但相反是附着在完整蛋白质上称为“膜靶蛋白”。在该提案中，特定的色谱修复蛋白混合物的一部分将用作亲和陷阱分离候选膜靶蛋白。这些蛋白质将被印迹到 PVDF 膜和部分测序。由此产生的信息将用于合成将用作免疫原的肽。肽特异性抗体预计可以标记发束的特定区域。 cDNA 文库将构建触手组织，然后用于转染细菌。将使用肽特异性抗体筛选重组克隆。从免疫阳性克隆中分离出单链 DNA，然后已测序。这项研究可能导致转导通道的隔离毛细胞中其他重要的膜蛋白。该信息将可能会导致更好地理解某些形式的分子基础耳聋和平衡障碍。